Although the differentiation stages in lineage committed hematopoietic cells are well characterized, the identify and regulation of the earliest progenitors produced from the first steps in differentiation of human lympho-hematopoietic stem cells (LHSC) remain unknown. The recent development of in vitro assays for primitive lymphoid progenitors now permits the direct study of events during early lymphoid commitment for human LHSC. We hypothesize that: (1) human lymphoid differentiation proceeds through two possible pathways, through Common Lymphoid Progenitors (CLP) (which lack myeloid and erythroid potential) and/or through B lympho-Myeloid Progenitors (BMP) (which lack myeloid and erythroid potential) and/or through B lympho-Myeloid Progenitors (BMP) (which lack T lymphoid and erythroid potential). These progenitor stages can be identified prospectively using immunophenotypic markers, and (2) the production (i.e. proliferation and differentiation) of murine and human lymphoid progenitor cells is coordinately regulated by three cytokine receptor signaling pathways: the Interleukin-7 Receptor alpha (IL-Ralpha)/Common gamma chain (gamma/c) complex; the IL- Ralpha/Thymic Stroma Derived Lymphopoietic Protein-Receptor (TSLP- R) complex; and the IL-3Ralpha/Common beta chain (beta/c) complex. The Specific Aims of this proposal are to (1) determine immunophenotypically the progenitor stages through which human LHSC differentiate into lymphoid cells, (2) characterize the differential gene expression in immunophenotypically and functionally defined populations of human LHSC and lymphoid progenitors, and (3) determine how lymphoid differentiation and proliferation from murine and human LHSC are regulated through the IL-7Ralpha and IL-3Ralpha mediated pathways. These studies will be performed using parallel and complementary in vitro and in vivo systems developed in our laboratories to study murine and human lympho-hematopoiesis.